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Abstract:

There are provided compounds and methods for the detection and treatment
of amyloid deposits and diseases and disorders characterized by amyloid
deposits including Alzheimer's disease and related amyloid-based
neurodegenerative disorders.

Claims:

1. A compound having the structure of Formula (I), ##STR00015## wherein
EDG is an electron donor group; πCE is a pi-conjugation element; and
WSG is a water soluble group.

3. The compound according to any of claims 1-2, wherein said
pi-conjugation element has the formula: -L1-(A1)q
L2-(A2)r-L3- or
-L1(A1)q-L4-A3-L2-(A2)r-L3-,
wherein q and r are independently 0 or 1; L1, L2, L3 and
L4 are independently a bond or a linking group having the formula:
##STR00016## wherein x is an integer from 1 to 50; A1, A2 and
A3 are independently R17-substituted or unsubstituted arylene,
or R17-substituted or unsubstituted heteroarylene; R17 is
halogen, --OR18, --NR19R20, R21-substituted or
unsubstituted alkyl, R21-substituted or unsubstituted heteroalkyl,
R21-substituted or unsubstituted cycloalkyl, R21-substituted or
unsubstituted heterocycloalkyl, R21-substituted or unsubstituted
aryl, or R21-substituted or unsubstituted heteroaryl; R18,
R19 and R.sup.°are independently hydrogen,
R21-substituted or unsubstituted alkyl, R21-substituted or
unsubstituted heteroalkyl, R21-substituted or unsubstituted
cycloalkyl, R21-substituted or unsubstituted heterocycloalkyl,
R21-substituted or unsubstituted aryl, or R21-substituted or
unsubstituted heteroaryl; R21 is halogen, --OR22,
--NR23R24, unsubstituted alkyl, unsubstituted heteroalkyl,
unsubstituted cycloalkyl, unsubstituted heterocycloalkyl, unsubstituted
aryl, or unsubstituted heteroaryl; and R22, R23 K and
R24 are independently hydrogen or unsubstituted alkyl.

4. The compound according to claim 3, wherein A1, A2 and
A3 are independently R21-substituted or unsubstituted
naphthylene, or R21-substituted or unsubstituted phenylene.

5. The compound according to claim 3, wherein x is an integer from 1 to
10.

6. The compound according to any of claims 1-5, wherein said water
soluble group is R25-substituted or unsubstituted alkyl,
R25-substituted or unsubstituted heteroalkyl, R25-substituted
or unsubstituted cycloalkyl, R25-substituted or unsubstituted
heterocycloalkyl, R25-substituted or unsubstituted aryl,
R25-substituted or unsubstituted heteroaryl; wherein R25 is
halogen, --OR26, --NR27R28, R29-substituted or
unsubstituted alkyl, R29-substituted or unsubstituted heteroalkyl,
R29-substituted or unsubstituted cycloalkyl, R29-substituted or
unsubstituted heterocycloalkyl, R29-substituted or unsubstituted
aryl, or R29-substituted or unsubstituted heteroaryl; R26,
R27 and R28 are independently hydrogen, R29-substituted or
unsubstituted alkyl, R29-substituted or unsubstituted heteroalkyl,
R29-substituted or unsubstituted cycloalkyl, R29-substituted or
unsubstituted heterocycloalkyl, R29-substituted or unsubstituted
aryl, or R29-substituted or unsubstituted heteroaryl, wherein
R27 and R28 are optionally joined together to form an
R29-substituted or unsubstituted heterocycloalkyl, or
R29-substituted or unsubstituted heteroaryl; R29 is halogen,
--OR30, --NR31R32, unsubstituted alkyl, unsubstituted
heteroalkyl, unsubstituted cycloalkyl, unsubstituted heterocycloalkyl,
unsubstituted aryl, or unsubstituted heteroaryl; and R30, R31
and R32 are independently hydrogen or unsubstituted alkyl.

7. The compound according to claim 6, wherein said water soluble group is
an ethylene glycol moiety having the formula: ##STR00017## wherein y is
an integer from 1 to 50.

8. The compound according to claim 6, wherein R29 is --OH.

9. The compound according to any of claims 1-8, said compound having the
structure: ##STR00018## ##STR00019##

10. A pharmaceutical composition comprising a compound according to any
of claims 1-9 and a pharmaceutically acceptable excipient.

11. A method of detecting an amyloid peptide, comprising contacting a
compound according to any one of claims 1-9 with an amyloid peptide
thereby forming a detectable amyloid complex, and detecting said
detectable amyloid complex.

12. The method according to claim 11, wherein said amyloid peptide is
Aβ peptide, prion protein, α-synuclein, or superoxide
dismutase.

13. The method according to claim 11, wherein said amyloid peptide forms
part of an amyloid.

14. A method of treating a disease characterized by an accumulation of
amyloids in a subject, comprising administering to a subject in need of
treatment an effective amount of a compound or pharmaceutical composition
according to any one of claims 1-10.

[0004] Compounds and methods for preventing or alleviating the symptoms of
amyloid-associated disease, for example but not limited to, neuronal
diseases and conditions associated with amyloid fibril or plaque
formation, have been provided in U.S. application Ser. No. 11/487,224,
filed Jul. 14, 2006, claiming priority to U.S. Prov. Appl. No.
60/699,728, filed Jul. 15, 2005, and U.S., Prov. Appl. No. 60/750,422,
filed on Dec. 13, 2005, and published as U.S. Published Appl. No.
2007/0066665, published Mar. 22, 2007, all of which are incorporated
herein by reference in their entireties and for all purposes. Compounds
and methods for the diagnosis and treatment of amyloid associated
diseases have been provided in International Appl. No. PCT/US2008/065410,
filed May 30, 2008, claiming priority to U.S. Prov. Appl. No. 60/940,968,
filed May 30, 2007, and published as PCT Publication No. WO 2008/15103,
on Dec. 11, 2008, all of which are incorporated herein by reference in
their entireties and for all purposes.

[0005] Each patent, published patent application, and reference cited
herein is hereby incorporated herein in its entirety and for all
purposes.

BRIEF SUMMARY OF THE INVENTION

[0006] In a first aspect, there is provided a compound having the
structure of Formula (I),

##STR00002##

[0007] The term "EDG" refers to an electron donor group, as known in the
art. The term "πCE" is a pi-conjugation element capable of being in
conjugation with the electron donor group. The term "WSG" refers to a
water soluble group.

[0008] In another aspect, there is provided a pharmaceutical composition.
The pharmaceutical composition includes a compound described herein and a
pharmaceutically acceptable excipient.

[0009] In another aspect, there is provided a method of detecting an
Aβ peptide. The method includes contacting a compound as described
herein with an Aβ peptide, and detecting a complex formed between
the compound with the Aβ peptide.

[0010] In another aspect, there is provided a method of treating a disease
or disorder characterized by an accumulation of amyloid deposits in the
brain. The method includes administering to a subject in need of
treatment an effective amount of a compound or pharmaceutical composition
as described herein.

[0018] FIG. 8 depicts the results of cytoxicity studies as described
herein. Legend: for each concentration of compound employed in the
cytoxicity assay, the % cell survival is plotted as a histogram in the
order (left to right): Cmpd 8a, 8b, 8c, 8d, 11 and 14, respectively.

DETAILED DESCRIPTION OF THE INVENTION

I. Definitions

[0019] The abbreviations used herein have their conventional meaning
within the chemical and biological arts. The chemical structures and
formulae set forth herein are constructed according to the standard rules
of chemical valency known in the chemical arts.

[0020] Where substituent groups are specified by their conventional
chemical formulae, written from left to right, they equally encompass the
chemically identical substituents that would result from writing the
structure from right to left, e.g., --CH2O-- is equivalent to
--OCH2--.

[0021] The term "alkyl," by itself or as part of another substituent,
means, unless otherwise stated, a straight (i.e. unbranched) or branched
chain, or combination thereof, which may be fully saturated, mono- or
polyunsaturated and can include di- and multivalent radicals, having the
number of carbon atoms designated (i.e. C1-C10 means one to ten
carbons). Examples of saturated hydrocarbon radicals include, but are not
limited to, groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl,
t-butyl, isobutyl, sec-butyl, (cyclohexyl)methyl, homologs and isomers
of, for example, n-pentyl, n-hexyl, n-heptyl, n-octyl, and the like. An
unsaturated alkyl group is one having one or more double bonds or triple
bonds. Examples of unsaturated alkyl groups include, but are not limited
to, vinyl, 2-propenyl, crotyl, 2-isopentenyl, 2-(butadienyl),
2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1- and 3-propynyl,
3-butynyl, and the higher homologs and isomers. An alkoxy is an alkyl
attached to the remainder of the molecule via an oxygen linker (--O--).

[0022] The term "alkylene" by itself or as part of another substituent
means a divalent radical derived from an alkyl, as exemplified, but not
limited, by --CH2CH2CH2CH2--, and further includes
those groups described below as "heteroalkylene." Typically, an alkyl (or
alkylene) group will have from 1 to 24 carbon atoms, with those groups
having 10 or fewer carbon atoms being preferred in the present invention.
A "lower alkyl" or "lower alkylene" is a shorter chain alkyl or alkylene
group, generally having eight or fewer carbon atoms.

[0023] The term "heteroalkyl," by itself or in combination with another
term, means, unless otherwise stated, a stable straight or branched
chain, or cyclic hydrocarbon radical, or combinations thereof, consisting
of at least one carbon atoms and at least one heteroatom selected from
the group consisting of O, N, P, Si and S, and wherein the nitrogen and
sulfur atoms may optionally be oxidized and the nitrogen heteroatom may
optionally be quaternized. The heteroatom(s) O, N, P and S and Si may be
placed at any interior position of the heteroalkyl group or at the
position at which the alkyl group is attached to the remainder of the
molecule. Examples include, but are not limited to,
--CH2--CH2--O--CH3, --CH2--CH2--NH--CH3,
--CH2--CH2--N(CH3)--CH3,
--CH2--S--CH2--CH3, --CH2--CH2--S(O)--CH3,
--CH2--CH2--S(O)2--CH3, --CH═CH--O--CH3,
--Si(CH3)3, --CH2--CH═N--OCH3,
--CH═CH--N(CH3)--CH3, O--CH3, --O--CH2--CH3,
and --CN. Up to two heteroatoms may be consecutive, such as, for example,
--CH,--NH--OCH3. Similarly, the term "heteroalkylene" by itself or
as part of another substituent means a divalent radical derived from
heteroalkyl, as exemplified, but not limited by,
--CH2--CH2--S--CH2--CH2-- and
--CH2--S--CH2--CH2--NH--CH2--. For heteroalkylene
groups, heteroatoms can also occupy either or both of the chain termini
(e.g., alkyleneoxy, alkylenedioxy, alkyleneamino, alkylenediamino, and
the like). Still further, for alkylene and heteroalkylene linking groups,
no orientation of the linking group is implied by the direction in which
the formula of the linking group is written. For example, the formula
--C(O )2R'-- represents both --C(O)2R'-- and --R'C(O)2--.
As described above, heteroalkyl groups, as used herein, include those
groups that are attached to the remainder of the molecule through a
heteroatom, such as --C(O)R', --C(O)NR', --NR'R'', --OR', --SR, and/or
--SO2R'. Where "heteroalkyl" is recited, followed by recitations of
specific heteroalkyl groups, such as --NR'R'' or the like, it will be
understood that the terms heteroalkyl and --NR'R'' are not redundant or
mutually exclusive. Rather, the specific heteroalkyl groups are recited
to add clarity. Thus, the term "heteroalkyl" should not be interpreted
herein as excluding specific heteroalkyl groups, such as --NR'R'' or the
like.

[0024] The terms "cycloalkyl" and "heterocycloalkyl," by themselves or in
combination with other terms, represent, unless otherwise stated, cyclic
versions of "alkyl" and "heteroalkyl", respectively. Additionally, for
heterocycloalkyl, a heteroatom can occupy the position at which the
heterocycle is attached to the remainder of the molecule. Examples of
cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl,
cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, and
the like. Examples of heterocycloalkyl include, but are not limited to,
1-(1,2,5,6-tetrahydropyridyl), 1-piperidinyl, 2-piperidinyl,
3-piperidinyl, 4-morpholinyl, 3-morpholinyl, tetrahydrofuran-2-yl,
tetrahydrofuran-3-yl, tetrahydrothien-2-yl, tetrahydrothien-3-yl,
1-piperazinyl, 2-piperazinyl, and the like. A "cycloalkylene" and a
"heterocycloalkylene," alone or as part of another substituent means a
divalent radical derived from a cycloalkyl and heterocycloalkyl,
respectively.

[0025] The terms "halo" or "halogen," by themselves or as part of another
substituent, mean, unless otherwise stated, a fluorine, chlorine,
bromine, or iodine atom. Additionally, terms such as "haloalkyl," are
meant to include monohaloalkyl and polyhaloalkyl. For example, the term
"halo(C1-C4)alkyl" is meant to include, but not be limited to,
fluoromethyl, difluoromethyl, trifluoromethyl, 2,2,2-trifluoroethyl,
4-chlorobutyl, 3-bromopropyl, and the like.

[0026] The term "acyl" means --C(O)R where R is a substituted or
unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted
or unsubstituted heteroalkyl, substituted or unsubstituted
heterocycloalkyl, substituted or unsubstituted aryl or substituted or
unsubstituted heteroaryl.

[0027] The term "aryl" means, unless otherwise stated, a polyunsaturated,
aromatic, hydrocarbon substituent which can be a single ring or multiple
rings (preferably from 1 to 3 rings) which are fused together (i.e. a
fused ring aryl) or linked covalently. A fused ring aryl refers to
multiple rings fused together wherein at least one of the fused rings is
an aryl ring. The term "heteroaryl" refers to aryl groups (or rings) that
contain from one to four heteroatoms selected from N, O, and S, wherein
the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen
atom(s) are optionally quaternized. Thus, the term "heteroaryl" includes
fused ring heteroaryl groups (i.e. multiple rings fused together wherein
at least one of the fused rings is a heteroaromatic ring). A 5,6-fused
ring heteroarylene refers to two rings fused together, wherein one ring
has 5 members and the other ring has 6 members, and wherein at least one
ring is a heteroaryl ring. Likewise, a 6,6-fused ring heteroarylene
refers to two rings fused together, wherein one ring has 6 members and
the other ring has 6 members, and wherein at least one ring is a
heteroaryl ring. And a 6,5-fused ring heteroarylene refers to two rings
fused together, wherein one ring has 6 members and the other ring has 5
members, and wherein at least one ring is a heteroaryl ring. A heteroaryl
group can be attached to the remainder of the molecule through a carbon
or heteroatom. Non-limiting examples of aryl and heteroaryl groups
include phenyl, 1-naphthyl, 2-naphthyl, 4-biphenyl, 1-pyrrolyl,
2-pyrrolyl, 3-pyrrolyl, 3-pyrazolyl, 2-imidazolyl, 4-imidazolyl,
pyrazinyl, 2-oxazolyl, 4-oxazolyl, 2-phenyl-4-oxazolyl, 5-oxazolyl,
3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, 2-thiazolyl, 4-thiazolyl,
5-thiazolyl, 2-furyl, 3-furyl, 2-thienyl, 3-thienyl, 2-pyridyl,
3-pyridyl, 4-pyridyl, 2-pyrimidyl, 4-pyrimidyl, 5-benzothiazolyl,
purinyl, 2-benzimidazolyl, 5-indolyl, 1-isoquinolyl, 5-isoquinolyl,
2-quinoxalinyl, 5-quinoxalinyl, 3-quinolyl, and 6-quinolyl. Substituents
for each of the above noted aryl and heteroaryl ring systems are selected
from the group of acceptable substituents described below. An "arylene"
and a "heteroarylene," alone or as part of another substituent means a
divalent radical derived from an aryl and heteroaryl, respectively.

[0028] For brevity, the term "aryl" when used in combination with other
terms (e.g., aryloxy, arylthioxy, arylalkyl) includes both aryl and
heteroaryl rings as defined above. Thus, the term "arylalkyl" is meant to
include those radicals in which an aryl group is attached to an alkyl
group (e.g., benzyl, phenethyl, pyridylmethyl and the like) including
those alkyl groups in which a carbon atom (e.g., a methylene group) has
been replaced by, for example, an oxygen atom (e.g., phenoxymethyl,
2-pyridyloxymethyl, 3-(1-naphthyloxy)propyl, and the like).

[0029] The term "oxo" as used herein means oxygen that is double bonded to
a carbon atom.

[0030] The term "alkylsulfonyl" as used herein means a moiety having the
formula --S(O2)--R', where R' is an alkyl group as defined above. R'
may have a specified number of carbons (e.g. "C1-C4
alkylsulfonyl").

[0031] Each of the above terms (e.g., "alkyl," "heteroalkyl," "aryl" and
"heteroaryl") are meant to include both substituted and unsubstituted
forms of the indicated radical. Preferred substituents for each type of
radical are provided below.

[0032] Substituents for the alkyl and heteroalkyl radicals (including
those groups often referred to as alkylene, alkenyl, heteroalkylene,
heteroalkenyl, alkynyl, cycloalkyl, heterocycloalkyl, cycloalkenyl, and
heterocycloalkenyl) can be one or more of a variety of groups selected
from, but not limited to: --OR', ═O, ═NR', ═N--OR', --NR'R'',
--SR', -halogen, --SiR'R''R''', --OC(O)R', --C(O)R', --CO2R',
--CONR'R'', --OC(O)NR'R'', --NR''C(O)R', --NR'--C(O)NR''R''',
--NR''C(O)2R', --NR--C(NR'R''R''')═NR'''',
--NR--C(NR'R'')═NR''', --S(O)R', --S(O)2R', --S(O)2NR'R'',
--NRSO2R', --CN and --NO2 in a number ranging from zero to
(2m'+1), where m' is the total number of carbon atoms in such radical.
R', R'', R''' and R'''' each preferably independently refer to hydrogen,
substituted or unsubstituted heteroalkyl, substituted or unsubstituted
cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or
unsubstituted aryl (e.g., aryl substituted with 1-3 halogens),
substituted or unsubstituted alkyl, alkoxy or thioalkoxy groups, or
arylalkyl groups. When a compound of the invention includes more than one
R group, for example, each of the R groups is independently selected as
are each R', R'', R''' and R'''' groups when more than one of these
groups is present. When R' and R'' are attached to the same nitrogen
atom, they can be combined with the nitrogen atom to form a 4-, 5-, 6-,
or 7-membered ring. For example, --NR'R'' is meant to include, but not be
limited to, 1-pyrrolidinyl and 4-morpholinyl. From the above discussion
of substituents, one of skill in the art will understand that the term
"alkyl" is meant to include groups including carbon atoms bound to groups
other than hydrogen groups, such as haloalkyl (e.g., --CF3 and
--CH2CF3) and acyl (e.g., --C(O)CH3, --C(O)CF3,
--C(O)CH2OCH3, and the like).

[0033] Similar to the substituents described for the alkyl radical,
substituents for the aryl and heteroaryl groups are varied and are
selected from, for example: halogen, --OR', --NR'R'', --SR', -halogen,
--SiR'R''R''', --OC(O)R', --C(O)R', --CO2R', --CONR'R'',
--OC(O)NR'R'', --NR''C(O)R', --NR'--C(O)NR''R''', --NR''C(O)2R',
--NR--C(NR'R''R'')═NR'''', --NR--C(NR'R'')═NR''', --S(O)R',
--S(O)2R', --S(O)2NR'R'', --NRSO2R', --CN and --NO2,
--R', --N3, --CH(Ph)2, fluoro(C1-C4)alkoxy, and
fluoro(C1-C4)alkyl, in a number ranging from zero to the total
number of open valences on the aromatic ring system; and where R', R'',
R''' and R'''' are preferably independently selected from hydrogen,
substituted or unsubstituted alkyl, substituted or unsubstituted
heteroalkyl, substituted or unsubstituted cycloalkyl, substituted or
unsubstituted heterocycloalkyl, substituted or unsubstituted aryl and
substituted or unsubstituted heteroaryl. When a compound of the invention
includes more than one R group, for example, each of the R groups is
independently selected as are each R', R'', R''' and R'''' groups when
more than one of these groups is present.

[0034] Two of the substituents on adjacent atoms of the aryl or heteroaryl
ring may optionally form a ring of the formula
-T-C(O)--(CRR')q--U--, wherein T and U are independently --NR--,
--O--, --CRR'--or a single bond, and q is an integer of from 0 to 3.
Alternatively, two of the substituents on adjacent atoms of the aryl or
heteroaryl ring may optionally be replaced with a substituent of the
formula -A-(CH2)r--B--, wherein A and B are independently
--CRR'--, --O--, --NR--, --S--, --S(O)--, --S(O)2--, --S(O)2NR'-- or
a single bond, and r is an integer of from 1 to 4. One of the single
bonds of the new ring so formed may optionally be replaced with a double
bond. Alternatively, two of the substituents on adjacent atoms of the
aryl or heteroaryl ring may optionally be replaced with a substituent of
the formula --(CRR')s--X'--(C''R''')d-, where s and d are
independently integers of from 0 to 3, and X' is --O--, --NR'--, --S--,
--S(O)--, --S(O)2--, or --S(O)2NR'--. The substituents R, R',
R'' and R''' are preferably independently selected from hydrogen,
substituted or unsubstituted alkyl, substituted or unsubstituted
cycloalkyl, substituted or unsubstituted heterocycloalkyl, substituted or
unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0035] As used herein, the term "heteroatom" or "ring heteroatom" is meant
to include oxygen (O), nitrogen (N), sulfur (S), phosphorus (P), and
silicon (Si).

[0036] A "substituent group," as used herein, means a group selected from
the following moieties:

[0043] A "size-limited substituent" or " size-limited substituent group,"
as used herein means a group selected from all of the substituents
described above for a "substituent group," wherein each substituted or
unsubstituted alkyl is a substituted or unsubstituted C1-C20
alkyl, each substituted or unsubstituted heteroalkyl is a substituted or
unsubstituted 2 to 20 membered heteroalkyl, each substituted or
unsubstituted cycloalkyl is a substituted or unsubstituted
C4-C8 cycloalkyl, and each substituted or unsubstituted
heterocycloalkyl is a substituted or unsubstituted 4 to 8 membered
heterocycloalkyl.

[0044] A "lower substituent" or " lower substituent group," as used herein
means a group selected from all of the substituents described above for a
"substituent group," wherein each substituted or unsubstituted alkyl is a
substituted or unsubstituted C1-C8 alkyl, each substituted or
unsubstituted heteroalkyl is a substituted or unsubstituted 2 to 8
membered heteroalkyl, each substituted or unsubstituted cycloalkyl is a
substituted or unsubstituted C5-C7 cycloalkyl, and each
substituted or unsubstituted heterocycloalkyl is a substituted or
unsubstituted 5 to 7 membered heterocycloalkyl.

[0045] The term "pharmaceutically acceptable salts" is meant to include
salts of the active compounds which are prepared with relatively nontoxic
acids or bases, depending on the particular substituents found on the
compounds described herein. When compounds of the present invention
contain relatively acidic functionalities, base addition salts can be
obtained by contacting the neutral form of such compounds with a
sufficient amount of the desired base, either neat or in a suitable inert
solvent. Examples of pharmaceutically acceptable base addition salts
include sodium, potassium, calcium, ammonium, organic amino, or magnesium
salt, or a similar salt. When compounds of the present invention contain
relatively basic functionalities, acid addition salts can be obtained by
contacting the neutral form of such compounds with a sufficient amount of
the desired acid, either neat or in a suitable inert solvent. Examples of
pharmaceutically acceptable acid addition salts include those derived
from inorganic acids like hydrochloric, hydrobromic, nitric, carbonic,
monohydrogencarbonic, phosphoric, monohydrogenphosphoric,
dihydrogenphosphoric, sulfuric, monohydrogensulfuric, hydriodic, or
phosphorous acids and the like, as well as the salts derived from
relatively nontoxic organic acids like acetic, propionic, isobutyric,
maleic, malonic, benzoic, succinic, suberic, fumaric, lactic, mandelic,
phthalic, benzenesulfonic, p-tolylsulfonic, citric, tartaric, oxalic,
methanesulfonic, and the like. Also included are salts of amino acids
such as arginate and the like, and salts of organic acids like glucuronic
or galactunoric acids and the like (see, for example, Berge et al.,
"Pharmaceutical Salts", Journal of Pharmaceutical Science, 1977, 66,
1-19). Certain specific compounds of the present invention contain both
basic and acidic functionalities that allow the compounds to be converted
into either base or acid addition salts.

[0046] Thus, the compounds of the present invention may exist as salts,
such as with pharmaceutically acceptable acids. The present invention
includes such salts. Examples of such salts include hydrochlorides,
hydrobromides, sulfates, methanesulfonates, nitrates, maleates, acetates,
citrates, fumarates, tartrates (e.g., (+)-tartrates, (-)-tartrates or
mixtures thereof including racemic mixtures), succinates, benzoates and
salts with amino acids such as glutamic acid. These salts may be prepared
by methods known to those skilled in the art.

[0047] The neutral forms of the compounds are preferably regenerated by
contacting the salt with a base or acid and isolating the parent compound
in the conventional manner. The parent form of the compound differs from
the various salt forms in certain physical properties, such as solubility
in polar solvents.

[0048] In addition to salt forms, the present invention provides
compounds, which are in a prodrug form. Prodrugs of the compounds
described herein are those compounds that readily undergo chemical
changes under physiological conditions to provide the compounds of the
present invention. Additionally, prodrugs can be converted to the
compounds of the present invention by chemical or biochemical methods in
an ex vivo environment. For example, prodrugs can be slowly converted to
the compounds of the present invention when placed in a transdermal patch
reservoir with a suitable enzyme or chemical reagent.

[0049] Certain compounds of the present invention can exist in unsolvated
forms as well as solvated forms, including hydrated forms. In general,
the solvated forms are equivalent to unsolvated forms and are encompassed
within the scope of the present invention. Certain compounds of the
present invention may exist in multiple crystalline or amorphous forms.
In general, all physical forms are equivalent for the uses contemplated
by the present invention and are intended to be within the scope of the
present invention.

[0050] Certain compounds of the present invention possess asymmetric
carbon atoms (optical centers) or double bonds; the racemates,
diastereomers, tautomers, geometric isomers and individual isomers are
encompassed within the scope of the present invention. The compounds of
the present invention do not include those which are known in the art to
be too unstable to synthesize and/or isolate.

[0051] The compounds of the present invention may also contain unnatural
proportions of atomic isotopes at one or more of the atoms that
constitute such compounds. For example, the compounds may be radiolabeled
with radioactive isotopes, such as for example tritium (3H),
iodine-125 (125I) or carbon-14 (14C). All isotopic variations
of the compounds of the present invention, whether radioactive or not,
are encompassed within the scope of the present invention.

[0052] Where a substituent of a compound provided herein is
"R-substituted" (e.g. R1-substituted), it is meant that the
substituent is substituted with one or more of the named R groups (e.g.
R1) as appropriate. In some embodiments, the substituent is
substituted with only one of the named R groups.

[0053] The terms "treating" or "treatment" refers to any indicia of
success in the treatment or amelioration of an injury, pathology or
condition, including any objective or subjective parameter such as
abatement; remission; diminishing of symptoms or making the injury,
pathology or condition more tolerable to the patient; slowing in the rate
of degeneration or decline; making the final point of degeneration less
debilitating; improving a patient's physical or mental well-being. The
treatment or amelioration of symptoms can be based on objective or
subjective parameters; including the results of a physical examination,
neuropsychiatric exams, and/or a psychiatric evaluation. For example, the
certain methods presented herein successfully treat cancer by decreasing
the incidence of cancer, in inhibiting its growth and or causing
remission of cancer.

[0054] An "effective amount" is an amount of a compound described herein
sufficient to contribute to the treatment, prevention, or reduction of a
symptom or symptoms of a disease, or to inhibit effects of an amyloid
relative to the absence of the compound. Where recited in reference to a
disease treatment, an "effective amount" may also be referred to as a
"therapeutically effective amount." A "reduction" of a symptom or
symptoms (and grammatical equivalents of this phrase) means decreasing of
the severity or frequency of the symptom(s), or elimination of the
symptom(s). A "prophylactically effective amount" of a drug is an amount
of a drug that, when administered to a subject, will have the intended
prophylactic effect, e.g., preventing or delaying the onset (or
reoccurrence) a disease, or reducing the likelihood of the onset (or
reoccurrence) of a disease or its symptoms. The full prophylactic effect
does not necessarily occur by administration of one dose, and may occur
only after administration of a series of doses. Thus, a prophylactically
effective amount may be administered in one or more administrations. An
"activity decreasing amount," as used herein, refers to an amount of
antagonist required to decrease the activity of an enzyme relative to the
absence of the antagonist. A "function disrupting amount," as used
herein, refers to the amount of antagonist required to disrupt the
function of an osteoclast or leukocyte relative to the absence of the
antagonist.

II. Molecular Rotor Motif Compounds

[0055] In one aspect, there is provided a compound having the structure of
Formula (I),

##STR00003##

[0056] The term "EDG" refers to an electron donor group, as known in the
art. The term "πCE" is a pi-conjugation element capable of being in
conjugation with the electron donor group. The term "WSG" refers to a
water soluble group.

[0063] In some embodiments, the EDG (Formula I) is substituted or
unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted
or unsubstituted heteroalkyl, substituted or unsubstituted
heterocycloalkyl, substituted or unsubstituted aryl, or substituted or
unsubstituted heteroaryl, --OR1, --NHC(O)R2, or
--NR3R4. In some embodiments, R1, R3 and R4 are
independently H, or substituted or unsubstituted alkyl; and R2 is
substituted or unsubstituted alkyl.

[0064] In some embodiments, the pi-conjugation element (Formula I) is
substituted or unsubstituted aryl, or substituted or unsubstituted
heteroaryl. In some embodiments, the pi-conjugation element is
unsubstituted aryl. In some embodiments, the pi-conjugation element is
substituted aryl.

[0065] In some embodiments, the water soluble group (Formula I) includes
ethylene glycol. In some embodiments, the water soluble group includes
polymeric ethylene glycol.

[0066] In some embodiments, a compound is provided which further includes
a detectable label. In some embodiments, the detectable label is a
radiolabel. In some embodiments, the detectable label is a fluorescent
label.

[0067] In some embodiments, each substituted group described herein in a
compound of Formula I is substituted with at least one substituent group.
More specifically, in some embodiments, each substituted alkyl,
substituted heteroalkyl, substituted cycloalkyl, substituted
heterocycloalkyl, substituted aryl, substituted heteroaryl, substituted
alkylene, substituted heteroalkylene, substituted or unsubstituted
cycloalkylene, substituted or unsubstituted heterocycloalkylene,
substituted or unsubstituted arylene, or substituted or unsubstituted
heteroarylene described herein in the compounds of Formula I is
substituted with at least one substituent group. In other embodiments, at
least one or all of these groups are substituted with at least one
size-limited substituent group. Alternatively, at least one or all of
these groups are substituted with at least one lower substituent group.

[0068] In other embodiments of the compounds of Formula I, each
substituted or unsubstituted alkyl is a substituted or unsubstituted
C1-C20 alkyl, each substituted or unsubstituted heteroalkyl is
a substituted or unsubstituted 2 to 20 membered heteroalkyl, each
substituted or unsubstituted cycloalkyl is a substituted or unsubstituted
C4-C8 cycloalkyl, each substituted or unsubstituted
heterocycloalkyl is a substituted or unsubstituted 4 to 8 membered
heterocycloalkyl, each substituted or unsubstituted alkylene is a
substituted or unsubstituted C1-C20 alkylene, each substituted
or unsubstituted heteroalkylene is a substituted or unsubstituted 2 to 20
membered heteroalkylene, each substituted or unsubstituted cycloalkylene
substituted or unsubstituted C4-C8 cycloalkylene, and each
substituted or unsubstituted heterocycloalkylene is a substituted or
unsubstituted 4 to 8 membered heterocycloalkylene.

[0069] Alternatively, each substituted or unsubstituted alkyl is a
substituted or unsubstituted C1-C8 alkyl, each substituted or
unsubstituted heteroalkyl is a substituted or unsubstituted 2 to 8
membered heteroalkyl, each substituted or unsubstituted cycloalkyl is a
substituted or unsubstituted C5-C7 cycloalkyl, each substituted
or unsubstituted heterocycloalkyl is a substituted or unsubstituted 5 to
7 membered heterocycloalkyl, each substituted or unsubstituted alkylene
is a substituted or unsubstituted C1-C8 alkylene, each
substituted or unsubstituted heteroalkylene is a substituted or
unsubstituted 2 to 8 membered heteroalkylene, each substituted or
unsubstituted cycloalkylene substituted or unsubstituted C5-C6
cycloalkylene, and each substituted or unsubstituted heterocycloalkylene
is a substituted or unsubstituted 5 to 7 membered heterocycloalkylene.

[0070] In some embodiments, a compound of Formula I is a compound within
the scope of Formula I set forth in Table 2 below:

[0071] Inspired by the structures of the currently used amyloid-binding
agents, we have evaluated the possibility to design new Aβ binding
fluorescent compounds useful as probes based on the molecular rotor
motif. We found that the molecular rotors bind to the aggregated AP
peptide with low micromolar affinity. We hypothesize that this binding is
a result of hydrophobic interactions between the rotor and the amyloid
peptide. This binding reduces the free volume around the rotor resulting
in an increased fluorescence emission. See R. O. Loutfy, B. A. Arnold, J.
Phys. Chem. 1982, 86:4205-4211; A. K. Doolittle, J. Appl. Phys. 1952,
23:236-239. A similar effect has been reported for the binding of
molecular rotors to actin, albumin and other proteins. See T. Lio, S.
Takahashi, S. Sawada, J. Biochem. 1993, 113:196-199; T. Iwaki, C.
Torigoe, M. Noji, M. Nakanishi, Biochem. 1993, 32:7589-7592. We have
demonstrated that these molecules can be readily synthesized and have no
significant cytotoxicity. In addition, we have shown that both the
physical properties and fluorescence profile of these fluorescent
compounds can be fine-tuned by modifying their chemical structure.
Notably, changes of the substitution at the electron donor group can
affect the intensity of fluorescence emission, while changes at the
π-system can affect the emission wavelength. These effects can be
implemented for the construction of multicolored dyes and can lead to
potential applications for in vitro and in vivo imaging. See C. J.
Sigurdson, K. Peter, R. Nilsson, S. Hornemann, G. Manco, M. Polymenidou,
P. Schwarz, M. Leclerc, P. Hammarstrom, K. Wuthrich, A. Aguzzi, Nat.
Methods 2007, 4,:023-1030. Interestingly, a recent report describes the
identification of CRANAD-2, a small molecule containing two
electron-donating groups connected simultaneously via π-conjugation to
a single difluoroboronate acceptor. See C. Ran, X. Xu, S. B. Raymond, B.
J. Ferrara, K. Neal, B. J. Bacskai, Z. Medarova, A. Moore, J. Am. Chem.
Soc. 2009, 131:15257-15261. This compound has a high affinity for Aβ
aggregates (Kd=38.0 nM) and suitable near-infrared fluorescence
properties for in vivo imaging, further validating our proposed concept
of exploring the molecular rotor motif for the development of new
amyloid-imaging compounds. These findings demonstrate that the D-π-A
motif of molecular rotors (Formula I) is a useful scaffold and represents
an important first step for the rational design of new diagnostic tools
for Alzheimer's disease and related amyloid-based neurodegenerative
disorders.

III. Pharmaceutical Compositions

[0072] In another aspect, the present invention provides pharmaceutical
compositions (i.e., formulations) comprising a compound described herein
in combination with a pharmaceutically acceptable excipient (e.g.,
carrier). The pharmaceutical compositions include optical isomers,
diastereomers, or pharmaceutically acceptable salts of the inhibitors
disclosed herein. For example, in some embodiments, the pharmaceutical
compositions include a compound of the present invention and citrate as a
pharmaceutically acceptable salt. The compound included in the
pharmaceutical composition may be covalently attached to a carrier
moiety, as described above. Alternatively, the compound included in the
pharmaceutical composition is not covalently linked to a carrier moiety.

[0073] A "pharmaceutically acceptable carrier," as used herein refers to
pharmaceutical excipients, for example, pharmaceutically,
physiologically, acceptable organic or inorganic carrier substances
suitable for enteral or parenteral application that do not deleteriously
react with the active agent. Suitable pharmaceutically acceptable
carriers include water, salt solutions (such as Ringer's solution),
alcohols, oils, gelatins, and carbohydrates such as lactose, amylose or
starch, fatty acid esters, hydroxymethycellulose, and polyvinyl
pyrrolidine. Such preparations can be sterilized and, if desired, mixed
with auxiliary agents such as lubricants, preservatives, stabilizers,
wetting agents, emulsifiers, salts for influencing osmotic pressure,
buffers, coloring, and/or aromatic substances and the like that do not
deleteriously react with the compounds of the invention.

[0074] The compounds of the invention can be administered alone or can be
coadministered to the subject. Coadministration is meant to include
simultaneous or sequential administration of the compounds individually
or in combination (more than one compound). The preparations can also be
combined, when desired, with other active substances (e.g. to reduce
metabolic degradation).

A. Formulations

[0075] The compounds can be prepared and administered in a wide variety of
oral, parenteral, and topical dosage forms. Thus, the compounds of the
present invention can be administered by injection (e.g. intravenously,
intramuscularly, intracutaneously, subcutaneously, intraduodenally, or
intraperitoneally). Also, the compounds described herein can be
administered by inhalation, for example, intranasally. Additionally, the
compounds of the present invention can be administered transdermally. It
is also envisioned that multiple routes of administration (e.g.,
intramuscular, oral, transdermal) can be used to administer the compounds
of the invention. Accordingly, the present invention also provides
pharmaceutical compositions comprising a pharmaceutically acceptable
carrier or excipient and one or more compounds of the invention.

[0076] For preparing pharmaceutical compositions from the compounds of the
present invention, pharmaceutically acceptable carriers can be either
solid or liquid. Solid form preparations include powders, tablets, pills,
capsules, cachets, suppositories, and dispersible granules. A solid
carrier can be one or more substance that may also act as diluents,
flavoring agents, binders, preservatives, tablet disintegrating agents,
or an encapsulating material.

[0077] In powders, the carrier is a finely divided solid in a mixture with
the finely divided active component. In tablets, the active component is
mixed with the carrier having the necessary binding properties in
suitable proportions and compacted in the shape and size desired.

[0078] The powders and tablets preferably contain from 5% to 70% of the
active compound. Suitable carriers are magnesium carbonate, magnesium
stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin,
tragacanth, methylcellulose, sodium carboxymethylcellulose, a low melting
wax, cocoa butter, and the like. The term "preparation" is intended to
include the formulation of the active compound with encapsulating
material as a carrier providing a capsule in which the active component
with or without other carriers, is surrounded by a carrier, which is thus
in association with it. Similarly, cachets and lozenges are included.
Tablets, powders, capsules, pills, cachets, and lozenges can be used as
solid dosage forms suitable for oral administration.

[0079] For preparing suppositories, a low melting wax, such as a mixture
of fatty acid glycerides or cocoa butter, is first melted and the active
component is dispersed homogeneously therein, as by stirring. The molten
homogeneous mixture is then poured into convenient sized molds, allowed
to cool, and thereby to solidify.

[0080] Liquid form preparations include solutions, suspensions, and
emulsions, for example, water or water/propylene glycol solutions. For
parenteral injection, liquid preparations can be formulated in solution
in aqueous polyethylene glycol solution.

[0081] When parenteral application is needed or desired, particularly
suitable admixtures for the compounds of the invention are injectable,
sterile solutions, preferably oily or aqueous solutions, as well as
suspensions, emulsions, or implants, including suppositories. In
particular, carriers for parenteral administration include aqueous
solutions of dextrose, saline, pure water, ethanol, glycerol, propylene
glycol, peanut oil, sesame oil, polyoxyethylene-block polymers, and the
like. Ampoules are convenient unit dosages. The compounds of the
invention can also be incorporated into liposomes or administered via
transdermal pumps or patches. Pharmaceutical admixtures suitable for use
in the present invention include those described, for example, in
Pharmaceutical Sciences (17th Ed., Mack Pub. Co., Easton, Pa.) and WO
96/05309, the teachings of both of which are hereby incorporated by
reference.

[0082] Aqueous solutions suitable for oral use can be prepared by
dissolving the active component in water and adding suitable colorants,
flavors, stabilizers, and thickening agents as desired. Aqueous
suspensions suitable for oral use can be made by dispersing the finely
divided active component in water with viscous material, such as natural
or synthetic gums, resins, methylcellulose, sodium
carboxymethylcellulose, and other well-known suspending agents.

[0083] Also included are solid form preparations that are intended to be
converted, shortly before use, to liquid form preparations for oral
administration. Such liquid forms include solutions, suspensions, and
emulsions. These preparations may contain, in addition to the active
component, colorants, flavors, stabilizers, buffers, artificial and
natural sweeteners, dispersants, thickeners, solubilizing agents, and the
like.

[0084] The pharmaceutical preparation is preferably in unit dosage form.
In such form the preparation is subdivided into unit doses containing
appropriate quantities of the active component. The unit dosage form can
be a packaged preparation, the package containing discrete quantities of
preparation, such as packeted tablets, capsules, and powders in vials or
ampoules. Also, the unit dosage form can be a capsule, tablet, cachet, or
lozenge itself, or it can be the appropriate number of any of these in
packaged form.

[0085] The quantity of active component in a unit dose preparation may be
varied or adjusted from 0.1 mg to 10000 mg, more typically 1.0 mg to 1000
mg, most typically 10 mg to 500 mg, according to the particular
application and the potency of the active component. The composition can,
if desired, also contain other compatible therapeutic agents.

[0086] Some compounds may have limited solubility in water and therefore
may require a surfactant or other appropriate co-solvent in the
composition. Such co-solvents include: Polysorbate 20, 60, and 80;
Pluronic F-68, F-84, and P-103; cyclodextrin; and polyoxyl 35 castor oil.
Such co-solvents are typically employed at a level between about 0.01%
and about 2% by weight.

[0087] Viscosity greater than that of simple aqueous solutions may be
desirable to decrease variability in dispensing the formulations, to
decrease physical separation of components of a suspension or emulsion of
formulation, and/or otherwise to improve the formulation. Such viscosity
building agents include, for example, polyvinyl alcohol, polyvinyl
pyrrolidone, methyl cellulose, hydroxy propyl methylcellulose,
hydroxyethyl cellulose, carboxymethyl cellulose, hydroxy propyl
cellulose, chondroitin sulfate and salts thereof, hyaluronic acid and
salts thereof, and combinations of the foregoing. Such agents are
typically employed at a level between about 0.01% and about 2% by weight.

[0088] The compositions of the present invention may additionally include
components to provide sustained release and/or comfort. Such components
include high molecular weight, anionic mucomimetic polymers, gelling
polysaccharides, and finely-divided drug carrier substrates. These
components are discussed in greater detail in U.S. Pat. Nos. 4,911,920;
5,403,841; 5,212,162; and 4,861,760. The entire contents of these patents
are incorporated herein by reference in their entirety for all purposes.

B. Effective Dosages

[0089] Pharmaceutical compositions provided by the present invention
include compositions wherein the active ingredient is contained in a
therapeutically effective amount, i.e., in an amount effective to achieve
its intended purpose. The actual amount effective for a particular
application will depend, inter alia, on the condition being treated. For
example, when administered in methods to treat cancer, such compositions
will contain an amount of active ingredient effective to achieve the
desired result (e.g. decreasing the number of cancer cells in a subject).

[0090] The dosage and frequency (single or multiple doses) of compound
administered can vary depending upon a variety of factors, including
route of administration; size, age, sex, health, body weight, body mass
index, and diet of the recipient; nature and extent of symptoms of the
disease being treated; presence of other diseases or other health-related
problems; kind of concurrent treatment; and complications from any
disease or treatment regimen. Other therapeutic regimens or agents can be
used in conjunction with the methods and compounds of the invention.

[0091] For any compound described herein, the therapeutically effective
amount can be initially determined from cell culture assays, as known in
the art.

[0092] Therapeutically effective amounts for use in humans may be
determined from animal models. For example, a dose for humans can be
formulated to achieve a concentration that has been found to be effective
in animals.

[0093] Dosages may be varied depending upon the requirements of the
patient and the compound being employed. The dose administered to a
patient, in the context of the present invention, should be sufficient to
affect a beneficial therapeutic response in the patient over time. The
size of the dose also will be determined by the existence, nature, and
extent of any adverse side effects. Generally, treatment is initiated
with smaller dosages, which are less than the optimum dose of the
compound. Thereafter, the dosage is increased by small increments until
the optimum effect under circumstances is reached. In one embodiment of
the invention, the dosage range is 0.001% to 10% w/v. In another
embodiment, the dosage range is 0.1% to 5% w/v.

[0094] Dosage amounts and intervals can be adjusted individually to
provide levels of the administered compound effective for the particular
clinical indication being treated. This will provide a therapeutic
regimen that is commensurate with the severity of the individual's
disease state.

[0095] Utilizing the teachings provided herein, an effective prophylactic
or therapeutic treatment regimen can be planned that does not cause
substantial toxicity and yet is entirely effective to treat the clinical
symptoms demonstrated by the particular patient. This planning should
involve the careful choice of active compound by considering factors such
as compound potency, relative bioavailability, patient body weight,
presence and severity of adverse side effects, preferred mode of
administration, and the toxicity profile of the selected agent.

C. Toxicity

[0096] The ratio between toxicity and therapeutic effect for a particular
compound is its therapeutic index and can be expressed as the ratio
between LD50 (the amount of compound lethal in 50% of the
population) and ED50 (the amount of compound effective in 50% of the
population). Compounds that exhibit high therapeutic indices are
preferred. Therapeutic index data obtained from cell culture assays
and/or animal studies can be used in formulating a range of dosages for
use in humans. The dosage of such compounds preferably lies within a
range of plasma concentrations that include the ED50 with little or
no toxicity. The dosage may vary within this range depending upon the
dosage form employed and the route of administration utilized. See, e.g.
Fingl et al., In THE PHARMACOLOGICAL BASIS OF THERAPEUTICS, Ch.1, p.1,
1975. The exact formulation, route of administration, and dosage can be
chosen by the individual physician in view of the patient's condition and
the particular method in which the compound is used.

IV. Methods of Use

[0097] In one aspect, there is provided a method of detecting an Aβ
peptide. The method includes contacting a compound as described herein
with an Aβ peptide, and detecting a complex formed between the
compound with the Aβ peptide, as described herein and known in the
art. The method of detection can employ spectroscopic (i.e., UV-visible,
fluorescence, and the like), radiographic, and other detection methods
known in the art.

[0098] In a further aspect, there is provided a method of treating a
disease or disorder characterized by an accumulation of amyloid deposits
in the brain. The method includes administering to a subject in need of
treatment a compound as described herein. In some embodiments, the
disease is Alzheimer's disease. The term "subject" as used herein refers
to a mammal to which a pharmaceutical composition or formulation is
administered. Exemplary subjects include humans, as well as veterinary
and laboratory animals such as horses, pigs, cattle, dogs, cats, rabbits,
rats, mice, and aquatic mammals.

V. Examples

[0099] General notes: All the reagents were obtained (Aldrich, Acros) at
highest commercial quality and used without further purification except
where noted. Air- and moisture-sensitive liquids and solutions were
transferred via syringe or stainless steel cannula. Organic solutions
were concentrated by rotary evaporation below 45° C. at
approximately 20 mmHg. All non-aqueous reactions were carried out under
anhydrous conditions. Yields refer to chromatographically and
spectroscopically (1H NMR, 13C NMR) homogeneous materials, unless
otherwise stated. Reactions were monitored by thin-layer chromatography
(TLC) carried out on 0.25 mm E. Merck silica gel plates (60E-254) and
visualized under UV light and/or developed by dipping in solutions of 10%
ethanolic phosphomolybdic acid (PMA) or p-anisaldehyde and applying heat.
E. Merck silica gel (60, particle size 0.040-0.063 mm) was used for flash
chromatography. Preparative thin-layer chromatography separations were
carried out on 0.25 or 0.50 mm E. Merck silica gel plates (60F-254). NMR
spectra were recorded on Varian Mercury 300 or 400 MHz instruments and
calibrated using the residual undeuterated solvent as an internal
reference. The following abbreviations were used to explain the
multiplicities: s=singlet, d=doublet, t=triplet, q=quartet, m=multiplet,
b=broad. High resolution mass spectra (HRMS) were recorded on a VG 7070
HS mass spectrometer under electron spray ionization (ESI) or electron
impact (EI) conditions. Fluorescence spectroscopy data were recorded on a
MD-5020 Photon Technology International Spectrophotometer at 25°
C.

[0100] General Procedure for the Preparation of Fluorescence Compounds. To
a round bottom flask containing a solution of aldehyde (5.0 mmol) and
2-(2-(2-mcthoxycthoxy) ethoxy)ethyl 2-cyanoacetate (5.5 mmol) in 20 ml of
THF was added 0.50 mmol of piperidine and the mixture was heated at
50° C. The reaction was monitored by TLC and was completed within
21 hours. The crude mixture was concentrated under reduced pressure and
the product was purified via flash chromatography (10-30% ethyl acetate
in hexane).

[0101] Example 1. Compound Synthesis and Analysis. Results of analysis of
compounds described herein are provided in Examples 1a-1m following.

[0116] In all cases, a 1.3-9.4 fold fluorescence intensity increase was
observed in the presence of aggregated Aβ, indicating that these
compounds bind to the peptide (Table 2). In most cases a modest
blue-shift (6-20 nm) was observed upon binding. Only in the case of the
naphthalene-based Cmpd 11 was a significant red shift of 76 nm observed
upon binding to preaggregated Aβ (FIGS. 1c, 1d). Interestingly, this
binding was accompanied with a 9.3 fold intensity increase. A similar
intensity increase has been observed with FDDNP and may be explained by
the ability of the naphthalene motif to create excimers upon binding to
its target. See E. D. Agdeppa, V. Kepe, J. Liu, S. Flores-Torres, N.
Satyamurthy, A. Petric, G. M. Cole, G. W. Small, S. C. Huang, J. R.
Barrio, J. Neurosci. 2001, 21:1-5; S. Abad, I. Vaya, M. C. Jimenez, U.
Pischel, M. A. Miranda, ChemPhysChem 2006, 7:2175-2183; C. Spies, R.
Gehrke J. Phys. Chem. A 2002, 106:5348-5352.

[0117] Cmpds 8a and 8b exhibited similar fluorescence characteristics
suggesting that addition of a methoxy group on the phenyl group does not
alter the binding properties of the compound as a probe. On the other
hand, it is worth noting that increasing the size of the alkyl groups of
the nitrogen leads to a significant increase in the fluorescence
intensity after binding (Table 2, 8a, 8c, 8d). This is likely a result of
the decreased rotational freedom of the molecules upon binding to the
aggregated forms of Aβ peptide. See W. Schuddeboom, S. A. Jonker, J.
M. Warman, U. Leinhos, W. Kuehnle, K. A. Zachariasse, J. Phys. Chem.
1992, 96 :10809-10819; Y. V. Il'chev, W. Kuehnle, K. A. Zachariasse, J.
Phys. Chem. 1998, 102 :5670-5680. Interestingly, no increase of
fluorescence intensity was observed upon mixing of these compounds with
monomeric Aβ peptide. This supports the notion that these compounds
bind selectively to aggregated forms of Aβ. The fluorescence profile
of 8d (excitation and emission) is shown in FIG. 1A and FIG. 1B.

[0120] The association of the synthesized compounds with aggregated
Aβ peptides was tested using a semi-quantitative ELISA based assay
developed by Yang and co-workers. See P. Inbar, J. Yang, Bioorg. Med.
Chem. Lett. 2006, 16:1076-1079; P. Inbar, C. Q. Li, S. A. Takayama, M. R.
Bautista, J. Yang, ChemBioChem 2006, 7:1563-1566; P. Inbar, M. R.
Bautista, S. A. Takayama, J. Yang, Anal. Chem. 2008, 80:3502-3506. The
assay is based on screening for molecules that inhibit the interaction of
the aggregated Aβ peptide with a monoclonal anti-Aβ IgG raised
against residues 1-17 of Aβ. Table 2 provides the concentrations of
the compounds corresponding to 50% inhibition (IC50) of the
IgG-Aβ interactions as well as the maximal percentage of the IgG's
inhibited from binding to the fibrils. All compounds exhibited IC50
values at μM levels, the lowest value being measured for 8b (IC50=1.17
μM). The maximum inhibition, a measure of the extent of surface
coating of the aggregated peptide by the compounds, was determined to be
between 40-98% (Table 2). See P. Inbar, J. Yang, Bioorg. Med. Chem. Lett.
2006, 16:1076-1079; P. Inbar, C. Q. Li, S. A. Takayama, M. R. Bautista,
J. Yang, ChemBioChem 2006, 7:1563-1566; P. Inbar, M. R. Bautista, S. A.
Takayama, J. Yang, Anal. Chem. 2008, 80:3502-3506. Comparison of these
data indicates that the surface coating increases by decreasing the size
of the compound or the extent of the π system. Specifically, while the
maximum inhibition is between 81-98% for the phenyl compounds, it
decreases to 58% for the longer naphthalene compound 11 and to 40% for
the more conjugated stilbene 19. Representative graphs for Cmpds 8d and
11 are shown in FIG. 3. Representative graphs for Cmpds 8a, 8b, 8c and 14
are shown in FIGS. 7A-D, respectively.

[0122] Example 4. Fluorescence Studies with Aggregated Aβ: Aggregated
Aβ peptide was prepared by dissolving Aβ(1-42) in PBS pH 7.4 to
a final concentration of 100 μM. This solution was magnetically
stirred at 1200 rpm for 3 days at room temperature. The 100 μM
Aβ(1-42) stock solution in PBS was aliquoted and frozen at
-80° C. for up to 4 weeks without noticeable change in its
property. 150 μL of pre-aggregated Aβ(1-42) was added to 2.85 mL
of compound to attain a final concentration of 5 μM Aβ(1-42) and
4 μM of compound. The solution was transferred to 3 mL cuvette and the
fluorescence measured at 25° C. As shown in FIG. 4, association of
compounds described herein with aggregated Aβ provides changes in
both excitation and emission spectra.

[0123] Example 5. Determination of Binding Constant: Pre-aggregated
Aβ(1-42) (5 μM final concentration) was mixed with various
concentrations of compounds described herein (10, 5, 2.5, 1.25 μM) in
PBS buffer (pH 7.4) and their fluorescence was measured. The negative
inverse of the x-intercept of the linear regression, that was drawn
between the double reciprocal of the fluorescence intensity maximum and
concentration of the compound, represents the compound binding constant
(Kd) to Aβ(1-42).

[0124] Example 6. Determination of Kd from Fluorescence Method. In
order to quantify the dissociation constants (Kd's) for the binding
of fluorescent compounds with aggregated (β-amyloid peptides, we
used the method described by LeVine (see H. LeVine III, Protein Sci.
1993, 2, 404-410). This method is similar to the method described by
Benesi-Hildebran (see C. Yang, L. Liu, T. W. Mu, Q. X. Guo, Anal. Sci.
2000, 16, 537-539). Here, the fluorescence of the compound was measured
with and without the addition of the aggregated peptides in solution. The
relative fluorescence enhancement of the compound upon binding to
aggregated (β-amyloid peptides was determined by taking the
difference between F (fluorescence after the addition of aggregated
peptides) and FO (fluorescence before the addition of aggregated
peptides).

[0125] In order to estimate the binding constant (Kd) for the
compound-Aβ complexes from the fluorescence studies, we made the
following assumptions:

[0126] 1. All compounds are completely in solution and free of any
significant competing binding process such as self-aggregation.

[0127] 2. The concentration of unbound compounds can be approximated as
close to the total concentration of the compounds.

[0128] 3. The binding sites in the aggregated Aβ peptides are not
completely occupied at the concentration of Aβ-binding compounds
used for the fluorescence studies (i.e., the experiments are carried out
under non-saturated binding conditions).

[0129] According to the Beer- Lambert law (see J. W. Robinson, "Atomic
spectroscopy", 1996), one can obtain two expressions that relate the
concentration of bound compound ([HG]), free compound ([G]), and free
amyloid peptides ([H]) with either 1) the measured fluorescence of the
compound in solution before the addition of the aggregated peptides
(FO), or 2) the measured fluorescence of the compound in the
presence of the amyloid peptides (F):

FO=εGl[GO] (1)

F=εHGl[HG]+εHl[H]+εGl[G] (2)

where [GO]=total concentration of compound

[0130] [HG]=compound- Aβ complex concentration

[0131] [HO]=total concentration of aggregated peptide

[0132] [II]=unbound aggregated peptide concentration.

[0133] εG=absorption coefficient of G

[0134] εHG=absorption coefficient of HG

[0135] εH=absorption coefficient of H

[0136] l=path length

[0137] Substituting into equation 1, and making the approximation that
εHGl[HG]+εGl[G]>>εHl[H], one
can arrive at a simplified expression for the relative fluorescence of
bound compound (ΔF):

ΔF=F-FQ=εHGl[HG]+εGl[G]-ε.s-
ub.Gl[G]-εGl[HG] (3)

or ΔF=Δεl[HG] (4)

where Δε=εHG-εG.

[0138] In order to obtain a relationship between the change in measured
fluorescence of the compound (ΔF) with the binding constant of the
compound to aggregated β-amyloid peptides (Kd's), we used the
standard equation for a binding isotherm to obtain a relationship between
[HG] and Kd:

[ HG ] = [ H O ] [ G ] K d + [ G ] ( 5 )
##EQU00001##

[0139] Combining equation 4 and 5, we obtained a relationship between
ΔF and Kd:

Δ F = [ H O ] [ G ] K d + [ G ]
Δ l ( 6 ) ##EQU00002##

[0140] In order to estimate the Kd of the compound bound to
aggregated Aβ peptides from the measured change in fluorescence, we
take the reciprocal of the equation 6 to give:

1 Δ F = K d Δ l [ H
O ] 1 [ G ] + 1 Δ l [ H O ]
( 7 ) ##EQU00003##

[0141] Equation 7 suggests that a double reciprocal plot of ΔF and
[G] should yield a straight line with x-intercept equal to -1/Kd.
FIG. 2 and FIG. 6 provide double reciprocal plots of the measured
fluorescence versus total concentration of compound [GO]. Assuming
that [G] can be approximated as close to [GO] (assumption 2), we can
obtain estimates for the Kd's of the compound-Aβ complexes from
the x-intercept of the linear fits of the data for each compound. The
estimated Kd's for all compounds are given in Table 2.

[0143] PBS buffer (various concentrations were obtained by diluting a
stock solution with PBS buffer) were incubated in the wells for 12 h.
Compounds that did not dissolve in PBS buffer were dissolved in DMSO and
diluted in PBS buffer to give a final solution of 5% DMSO in PBS buffer.
The excess solutions were then removed and all wells were blocked for 30
min by adding 300 μL of a 1% (w/v) solution of bovine serum albumin in
PBS buffer (BSA/PBS). On occasion, an additional blocking step was
performed prior to incubation with solutions of small molecules. The
blocking solution was discarded and the wells were washed once with 300
μL of PBS buffer. Wells were incubated for 1 h with 50 μL of a 1.1
nM solution (in 1% BSA/PBS, dilution 1:6000) of anti-Aβ IgG (clone
6E10, monoclonal, mouse), followed by removal of the solution. The wells
were washed twice with 300 μL of PBS buffer and incubated for 60 min
with 50 μL of the secondary IgG (anti- mouse IgG H+L, polyclonal,
rabbit) conjugated with alkaline phosphatase (6.8 nM in 1% BSA/PBS,
dilution 1:1000). The solution was discarded, and the wells were washed
twice with 300 μL PBS buffer. Bound secondary IgGs were detected by
the addition of 50 μL of a p-nitrophenyl phosphate solution (2.7 mM,
in 100 mM diethanol amine/0.5 mM magnesium chloride, pH 9.8). Absorbance
intensities were determined at 405 nm using a UV-vis spectroscopic plate
reader (Sprectramax 190, Molecular Devices, Sunnyvale, Calif.). Each run
was performed five times and averaged. Error bars represent standard
deviations. Graphs were plotted and fitted with the sigmoid curve
fitting.

[0144] Example 8. Fluorescence Studies with Monomeric Aβ. Aβ
(Biopeptide, Inc.) was initially solubilized in hexafluoroisopropanol at
1 mM concentration, vortexed, sonicated and vortexed. The vial was
covered in foil and was incubated for 21 hours at 25° C. on a
shaker, with 3 times of vortexing throughout the incubation period. The
solution was sonicated and vortexed again then diluted with cold nanopure
water (2:1 H2O:HFIP), fractionated in desired amounts into small
glass vials, and immediately frozen in a CO2/acetone bath. Each
fraction was covered with parafilm that was punctured to allow solvent
vapors to escape. The fractions were lyophilized for 2 days to obtain
monomeric Aβ (91% monomer by 12% Tris-bis PAGE gel analysis). 1.8
μL (8.42 μM) of this monomeric Aβ(1-42) was added to 3 μL
of 4 μM concentration of small molecules that was prepared by
dissolving in PBS buffer pH 7.4 to attain a final concentration of 5
μM of Aβ(1-42) and 4 μM of the compound. The solution was
transferred to 3 mL cuvettes and the fluorescence was measured at
25° C.

[0145] Example 9. Evaluation of Rigid Rotors for Cytotoxic Activity
Against SHSY-5Y human neuroblastoma cells (MTT assay): SHSY-5Y human
neuroblastoma cells, MTT
(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) cell
proliferation kit, Eagle's Minimum Essential Medium (EMEM), Ham's F12
nutrient mixture, and Fetal Bovine Serum (FBS) were all purchased from
ATCC (Manassas, Va.). Briefly, SH-SY5Y cells (in 1:1 EMEM:Ham's F-12 with
10% FBS) were seeded on 96-well plates at a density of 5×104
cells/well. Plates were incubated overnight (in a humidified atmosphere
of 95% air, 5% CO2, at 37° C.) to promote attachment of cells
to the wells. Cells were then treated with various concentrations of
compound 8a, 8b, 8c, 8d, 11, or 14 and incubated for 24 hours (humidified
atmosphere of 95% air, 5% CO2, at 37° C.). MTT reagent (20
μL) was added to the medium and incubated for additional 4 hours.
After incubation, 100 μL of detergent reagent was added and the plates
were covered with aluminum foil and left at room temperature overnight.
The amount of solubilized MTT formazan was measured by spectrophotometric
absorbance at 570 nm (Spectramax 190, Molecular Devices, Sunnyvale,
Calif.). MTT assay was not performed on compound 19 due to its poor
solubility in aqueous media. All data are presented as the mean±S.D,
N=3 for each concentration. The Student's t-test was employed for all
analyses. A p-value of <0.05 was considered statistically significant
compared to control cells. As shown in FIG. 8, all compounds showed
little or no cytotoxicity against human neuroblastoma cells at
concentrations up to 100 μM. These properties represent significant
advantages for further in vivo evaluation.